The subunits of spinach chloroplast CF, separated in SDS on hydroxylapatite and gel filtration columns, will be examined in detail for the location of free -SH gruops and of the half-cystines involved in disulfide bonds. Current indications of microheterogeneity with respect to the latter function in beta and gamma subunits will be tested more critically. Varieties of CF1 to be examined include the enzyme from untreated chloroplasts, from those with ATPase activated by illumination plus DTT, and perhaps from those with CF1 oxidized and inactivated with iodosobenzoate in the light. Dr. R. Bradshaw at the University of Washington will attempt to determine the amino acid sequence of the smallest, epsilon, subunit. We also hope to have his collaboration in finding the N-terminal sequence of each of the subunits, then of the whole enzyme, to assess their relative stoichiometry in the native enzyme. Stoichiometry will also be measured using uniformly-labeled CF obtained from plants grown in CO214. New reactive chemicals will be screened for possible energy-dependent and/or active site inhibitions of CF1. Dimethylsuberimidate is under examination as a reagent which might prevent conformational movements and inhibit phosphorylation for this reason without preventing its "hole-plugging" action for trans-membrane protons. Further attempts will be made to obtain reconstitutable membranes with CF1 either removed completely, or specifically and completely inactivated, in order to have a valid assay for catalytic activity in photophosphorylation of newly added CF1. BIBLIOGRAPHIC REFERENCES: Jagendorf, A.T. (1975) Fed. Proc. 34, 1718-1722. Chloroplast membranes and coupling factor conformations.